1. Field of the Invention
The present invention is generally related to culturing of cells and more particularly to a roller bottle suitable for trans-membrane co-culturing of cells.
2. Description of Related Information
Culturing of cells of various types has become a routine process in many laboratories. Cells are grown to harvest compounds, to test for various sensitivities to potentially toxic compounds and even to provide tissue for grafts. More recently, cells of different types have been co-cultured on opposite sides of a microporous membrane for the study of interactions between one type of cell and another. The function of the microporous membrane is to prevent physical contact of one cell type with the other cell type while allowing a chemical communication between the cell types using the cellular metabolites and cellular products soluble in the aqueous media. This procedure is called "trans-membrane co-culture."
Representative references to the co-culture of cells include Mangum et al., In Vitro Cell Dev. Biol. 26:1135-1143 (Dec., 1990) in which the authors describe a "Co-Culture of Primary Pulmonary Cells to Model Alveolar Injury and Translocation of Protein"; and Madara et al. in J. Tissue Cult. Method. 14:209-216, (1992) report "A Simple Approach to Measurement of Electrical Parameters of Cultured Epithelial Monolayers: Use in Assessing Neutrophil-Epithelial Interactions." Both of these papers and the references therein report a technique for growing cells of two different types on opposite sides of a suspended microporous membrane to study the interactions between the cells. Madara et al. provide a detailed report of a trans-membrane co-culture study. Madara et al. also describe modifications of a commercial cell culture insert by adhesively bonding a machined polycarbonate ring onto the underside of the insert to facilitate the growth of monolayers on both sides of the membrane. These modifications of an existing product described by Madara et al., provided them with a way to conduct a small scale experiment on interactions of two types of cells, but the equipment developed by Madara et al. is not suitable for larger scale studies.
As the technique of trans-membrane co-culture of cells becomes of greater interest, there is a need to grow larger populations of co-cultured cells than can easily be handled using small (1-5 cm.) well insert apparatus. One example demonstrating the need for larger scale co-culture is that some types of cellular metabolite products are secreted by one type of cells only in the presence of other types of cells. These metabolite products are produced in very small quantities in a co-culture of the cells. The effect of these metabolites from one cell type on another cell type may be observed in the small well insert apparatus, but in order to isolate sufficient quantities of these metabolites for detailed characterization, it is desirable to grow larger populations of the co-culture cells.
In scale up of cell monocultures, the containers used in laboratory settings for cell culturing are commonly known as "roller bottles." These roller bottles are generally cylindrical and are adapted to rotate about their longitudinal axis when placed on a laboratory roller apparatus. The laboratory roller apparatus is often adapted to fit within chambers for controlling the temperature and atmosphere. The bottles have evolved into standard sizes of about 0.85, 1.75 and 3.5 liters capacity and about 11-12 cm. in diameter because of the constraints of the roller apparatus.
The most common application for these bottles is to be charged with a few hundred milliliters (ml) of aqueous growth medium containing a suspension of the cells to be cultured. The bottles are placed on their sides on a laboratory roller apparatus, generally in a temperature and atmosphere controlled environment, so that a longitudinal portion of the inside surface of the bottle is always wet by the growth medium, and rotated slowly (about 1-2 rpm) about their axis for several days. During this time, the cells grow and utilize the nutrients present in the growth medium, generally by attaching themselves to the surface of the bottle. In many applications, it is necessary to drain and recharge the growth medium during the course of the growth cycle, either to harvest by-products, remove waste or replenish nutrients.
There is a need for devices similar to mono-culture roller bottles for scaling up of trans-membrane co-culture systems. Such a device and a method for its use are described below.